ABSTRACT
Most drugs need to interact with cell membrane to reach the biological target, so that membrane affinity assay is an important early screening step in drug discovery. However, at present, the traditional oil-water distribution method is still used, a new, simple and accurate method for membrane affinity assay is urgently needed. In this study, according to the colorimetric principle, a new assay model based on polydiacetylene vesicles was optimized through a series of experiments including different concentrations of vesicle solution, temperature, or pH reaction environment. On this basis, tetracaine hydrochloride, 2-methylimidazole and histamine were used as model drugs to measure the membrane affinity constants and verify the between-batch precision of the optimized assay model (relative standard deviation less than 5%). In addition, polydiacetylene vesicles were stable for up to 180 days, demonstrating the potential application of the assay model. This strategy is simple, stable, reliable, with high reproducibility, low cost and easy to promote, which provided a new tool and a new direction for the high-throughput assay of membrane affinity.
ABSTRACT
The study was designed to explore the active components and mechanism of Kai Xin San in the treatment of Alzheimer's disease (AD) based on network pharmacology. All targets related to AD were researched in the Traditional Chinese Medicine Systems Pharmacology Database and Analysis Platform (TCMSP) and Therapeutic Targets Database (TTD). The common targets obtained by two databases were determined as candidate proteins involved in AD. All active components related to Kai Xin San were researched from ADME (absorption, distribution, metabolism and excretion). PharmMapper was used to obtain the primary candidate targets of Kai Xin San. The corresponding gene name of each target protein was obtained from the UniProt database and selected human target proteins. Finally, the target proteins related to AD by Kai Xin San were acquired; Cytoscape 3.5.1 was used to construct the topology analysis for the active ingredient-AD target interaction network of Kai Xin San. According to STRING database and DAVID annotation databases, Gene Ontology enrichment analysis and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis of the targets was performed. The network pharmacology analysis results were verified by Discovery Studio molecular docking software. There were 31 components meeting the conditions of ADME and 8 targets relating to AD. Thirteen kinds of biological process, 7 related to molecular function and 11 related to cellar components, were included in 31 GO entries. There were 5 KEGG pathways, involving the calcium signaling pathway and PI3K-Akt signaling pathway. The docking results of Discovery Studio showed that active ingredients of Kai Xin San and the positive controls all have good binding activity with important targets. In conclusion, the Kai Xin San as applied for treating AD has the advantages of multi-components and targets, to investigate the active components and mechanism of Kai Xin San for treating AD based on network pharmacology to eludicate possible studies of the mechanisms of action.
ABSTRACT
BACKGROUND: The 3D printed polylactic-co-glycolicacid/nano-hydroxyapatite (PLGA/nHA) scaffold carrying human recombinant bone morphogenetic protein 2 (rhBMP-2)/chitosan (CS) sustained release tissue-engineering bone has good biological activity, mechanical properties, and biological activity of its controlled release rhBMP-2. OBJECTIVE: To investigate the repair of mandibular defects with PLGA/nHA scaffold/rhBMP-2/CS sustained release tissue-engineering bone manufactured using 3D bionic printing technology. METHODS: Animal models of bilateral critical mandibular bone defects were established in 27 New Zealand white rabbits, followed by implantation of a 3D-printed PLGA/nHA scaffold/rhBMP-2/CS sustained release tissue-engineering bone on one side (experimental side), and a 3D-printed PLGA/nHA scaffold on the other side (control side). Mandibular specimens were harvested at postoperative days 30, 60 and 90 to carry out cone-beam CT, Micro CT, histological and immunohistochemical examinations. RESULTS AND CONCLUSION: The results from micro-CT analysis revealed that the volume of newly formed bone volume and the amount of bone trabeculae on the experimental side were significantly higher than those on the control side at different postoperative time points (P ＜ 0.05). The results from cone-beam CT examination showed that at 90 postoperative days, bone density of the bone defect on the experimental side was close to that of the surrounding bone, new bone tissues were full of the original bone defect area, and the trabecular bone arranged regularly, while on the control side, worm-eaten discontinuous low-density osteoid tissues were visible in the bone defect area. Osteogenesis on the experimental side was better than that on the control side. Histological findings demonstrated that on the experimental side, a large amount of mature lamellae were detected in the bone defect area, with well-arranged trabecular bones and abundant capillaries, and moreover, the scaffold material had been completely absorbed. However, low-density, loose-meshed, irregular braided bone tissues with rare capillaries were observed on the control side, and the scaffold material had been mostly absorbed. Immunohistochemical findings indicated that the osteocalcin-dyed area on the experimental side was significantly larger than that of the control side at postoperative 90 days. To conclude, 3D-printed PLGA/nHA scaffold/rhBMP-2/CS sustained release tissue-engineering bone is favorable for the repair and reconstruction of experimental mandibular defects in rabbits.